Abstract:

Objective To discuss the change of sensitivity of the non-small-cell lung cancer （NSCLC） A549 cells to doxorubicin （Dox） after silencing forkhead protein 3 （FOXP3） gene，and to clarify its mechanism involved in Dox resistance. Methods The human NSCLC A549 cells were transfected with FOXP3 small interfering RNA（siRNA ）by lipofectamine method. The cells were divided into blank control group （without transfection）， si-NC group （transfected with control-siRNA）， and si-FOXP3 group （transfected with FOXP3-siRNA）. Western blotting and immunofluorescence methods were used to detect the expression levels of FOXP3 protein in the A549 cells in various groups；the proliferation activities and half-maximal inhibitory concentration （IC₅₀） values of the A549 cells in various groups were detected by CCK-8 method. The A549 cells were treated with 0， 10， and 20 μmol·L^－1 DAPT， and regarded as 0， 10， and 20 μmol·L^－1 DAPT groups， respectively. Additionally， the A549 cells were treated with 0 μmol·L^－1 DAPT， 1.0 mg·L^－1 Dox， and 1.0 mg·L^－1 Dox combined with 10 μmol·L^－1 DAPT， and regarded as 0 μmol·L^－1 DAPT， 1.0 mg·L^－1 Dox， and 1.0 mg·L^－1 Dox combined with 10 μmol·L^－1 DAPT groups， respectively. Western blotting method was used to detect the expression levels of Notch1，Hes1， and FOXP3 proteins in the A549 cells in various groups； the IC₅₀ values and expression levels of Notch1， Hes1， and FOXP3 proteins in the A549 cells in 0， 10， and 20 μmol·L^－1 DAPT groups were detected by CCK-8 and Western blotting methods；the expression levels of FOXP3， P-glycoprotein（P-gp）， Notch1， and Hes1 proteins in the A549 cells in 0 μmol·L^－1 DAPT， 1.0 mg·L^－1 Dox， and 1.0 mg·L^－1 Dox combined with 10 μmol·L^－1 DAPT groups were detected by Western blotting method. Results Compared with blank control and si-NC groups， the expression level of FOXP3 protein in the A549 cells in si-FOXP3 group was significantly decreased （P<0.01）， the proliferative activity and IC₅₀ value were decreased （P<0.05 or P<0.01），and the expression levels of Notch1，Hes1 and FOXP3 proteins were significantly decreased （P<0.01）. Compared with 0 μmol·L^－1 DAPT group， the expression levels of Notch1， Hes1， and FOXP3 proteins in the A549 cells in 10 and 20 μmol·L^－1 DAPT groups were significantly decreased （P<0.01）. Compared with 0 μmol·L^－1 DAPT group， the expression levels of FOXP3， P-gp， Notch1， and Hes1 proteins in the A549 cells in 1.0 mg·L^－1 Dox group were significantly increased （P<0.01）. Compared with 1.0 mg·L^－1 Dox group， the expression levels of FOXP3， P-gp， Notch1， and Hes1 proteins in the A549 cells in 1.0 mg·L^－1 Dox combined with 10 μmol·L^－1 DAPT group were significantly decreased （P<0.01）. Conclusion Silencing FOXP3 can enhance the sensitivity of the NSCLC cells to Dox， and its mechanism is related to the inhibition of the Notch1/Hes1 signaling pathway.

Key words: Cancer，non small-cell lung, Forkhead box protein 3, Notch1, Doxorubicin, Drug sensitivity